The cAMP signaling system regulates LHbeta gene expression: roles of early growth response protein-1, SP1 and steroidogenic factor-1

Abstract

Expression of the gonadotropin genes has been shown to be modulated by pharmacological or physiological activators of both the protein kinase C (PKC) and the cAMP second messenger signaling pathways. Over the past few years, a substantial amount of progress has been made in the identification and characterization of the transcription factors and cognate cis-elements which mediate the PKC response in the LH -subunit (LH ) gene. In contrast, little is known regarding the molecular mechanisms which mediate cAMP-mediated regulation of this gene. Using pituitary cell lines, we now demonstrate that rat LH gene promoter activity is stimulated following activation of the cAMP system by the adenylate cyclase activating agent, forskolin, or by the peptide, pituitary adenylate cyclase-activating peptide. The forskolin response was eliminated with mutation of a previously identified 3′ cis-acting element for the early growth response protein-1 (Egr-1) when evaluated in the context of region −207/+5 of the LH gene. Activation of the cAMP system increased Egr-1 gene promoter activity, Egr-1 protein levels and Egr-1 binding to the LH gene promoter, supporting the role of this transcription factor in mediating the cAMP response. Analysis of a longer LH promoter construct (−797/+5) revealed additional contribution by upstream Sp1 DNA-regulatory regions. Of interest, forskolin-induced stimulation of LH gene promoter activity was observed to increase synergistically with introduction of the transcription factor, steroidogenic factor-1 (SF-1). Although SF-1 is a critical mediator of the cAMP response in other genes, mutation of the SF-1 DNA-binding sites in the rat LH gene did not alter the forskolin response nor did forskolin increase SF-1 protein levels in a gonadotrope cell line. In a further set of experiments, it was determined that forskolin-responsiveness was maintained following mutation of the previously defined homeobox-binding element at position -100. We conclude that both Egr-1 and Sp1 contribute to cAMP-dependent transcription of the rat LH gene promoter. While SF-1 does not act independently to mediate the cAMP/PKA response, SF-1 is important for magnification of this response.

Gonadotropin-releasing hormone and the control of gonadotrope function

“…GnRH can also induce phospholipases D and A2, as well as a delayed production of cAMP however, the mechanisms involved remain obscure. Cyclic AMP has been shown to be capable of inducing increased expression of LH subunit genes and the release of newly synthesised LH [32,33] as well as an increased expression of the GnRH receptor [34] and NOS I [35,36] genes.…”

The GnRH receptor and the response of gonadotrope cells to GnRH pulse frequency code. A story of an atypical adaptation of cell function relying on a lack of receptor homologous desensitization.

“…Recent data suggest the contribution of novel PKC isoforms in this process [20] in addition to the direct coupling through Gαs [21]. Cyclic AMP stimulates the expression of LH subunit genes as well as the release of newly synthesised LH [22][23][24]. Collectively, these data indicate that GnRH induces a complex network of intracellular signaling pathways with different kinetics to activate or inhibit gonadotrope functions.…”

“…PKC, MAPK as well as the cAMP/PKA pathway [24]. Such a complex signaling could reflect redundancy or may be necessary for a coordinate regulation of cell functions.…”

“…This strengthens the idea that the cAMP/PKA signaling pathway would be preferentially operative under sustained GnRH stimulation. Moreover, cAMP can indirectly induce NAB2 since it stimulates Egr-1 promoter activity [24]. All the PKC, MAPK and cAMP/PKA transduction systems can thus potentially either activate or inhibit the LHβ gene depending on the modalities of GnRH stimulation.…”

“…Several of these genes are members of the GnRHR signaling network. These include, notably, the GnRHR itself [10,26], NOS I [27,28], PKA [29] and PKC [30] isoforms as well as transcription factors such as ATF3 [31], NUR77 [32] and EGR-1 [24], or autocrine/ paracrine factors such as activin and follistatin. Regulation of these genes by GnRH appeared to be mediated through two or more signaling pathways.…”